SUNDAY, DECEMBER 4- Late Abstracts 1 - Molecular Biology of the ...

SUNDAY
We have established an automated image-processing pipeline that allows us to quantitatively
monitor lamina disassembly in live HeLa cells. Using kinase inhibitors, we could show that both
CDK1 and PKCs significantly contribute to efficient Lamin B1 disassembly in HeLa cells.
Similarly, overexpression of Lamin B1 reporters with mutated CDK1 or PKC consensus
phosphorylation sites also impeded Lamin B1 disassembly. Since the role of PKC-dependent
pathways during mitosis and their effects on lamina disassembly are poorly understood, we
focused on elucidating this process in more detail. Using RNAi we found that specific PKC
isoenzymes that require diacylglycerol (DAG) for their activation trigger rate-limiting steps during
mitotic Lamin B1 disassembly in our in vivo assay.
Interestingly, we recently showed that the enzyme Lipin, which produces DAG by
dephosphorylation of phosphatidate, is essential for Lamin disassembly in the nematode C.
elegans (Gorjánácz and Mattaj, 2009). This suggested that Lipin might act upstream of DAGdependent
PKCs. To investigate this possibility we used RNAi to deplete the three redundant
human Lipin enzymes and monitored Lamin B1 disassembly. Strikingly, Lipin depletion inhibited
Lamin B1 disassembly to a similar extent as did PKC depletion. Furthermore, the effect of Lipin
RNAi could be rescued by the addition of a DAG analogue.
This supports a model where Lipins function in a lipid-mediated manner to activate a PKCdependent
pathway of mitotic Lamin disassembly.
Gorjánácz, M., and Mattaj, I.W. (2009). Lipin is required for efficient breakdown of the nuclear
envelope in Caenorhabditis elegans. J Cell Sci 122, 1963-1969.
Signal Transduction and Signaling Networks I
2026
Negative regulation of Notch1 signaling by Serum- and glucocorticoid-inducible kinase 1.
J-H. Yoon 1 , E-J. Ann 1 , J-S. Ahn 1 , H-S. Park 1 ; 1 Hormone Research Center, School of Biological
Sciences and Technology, Chonnam National University, Gwangju, Korea
Notch is a transmembrane protein that acts as a transcriptional factor in the Notch signaling
pathway for cell survival, cell death, and cell differentiation. However, little is known about
mechanisms and regulators that are responsible for attenuating the Notch signaling pathway
and protein stability. Here, we report that SGK1 remarkably reduced the protein stability of
Notch1 through Fbw7. The protein level and transcriptional activity of the Notch1 intracellular
domain were higher in SGK1 null cells than in SGK1 wild-type cells. Notch1-IC was able to form
a trimeric complex with Fbw7 and SGK1, thereby SGK1 enhanced the protein degradation of
Notch1-IC via a Fbw7-dependent proteasomal pathway. Furthermore, activated SGK1
phosphorylated Fbw7 at Ser227, an effect inducing Notch1-IC protein degradation and
ubiquitination. Moreover, accumulated dexamethasone-induced SGK1 facilitated the
degradation of Notch1-IC through phosphorylation of Fbw7. Altogether our results suggest that
SGK1 inhibits the Notch1 signaling pathway via phosphorylation of Fbw7.